AMG 191 depletes human HSPCs engrafted in NSG mice and second human donor engraftment level correlates with AMG 191 dose administered. (A) Representative FACS plots showing hCD34⁺hCD117⁺ cell population gated on hCD45⁺ cells present in BM aspirates obtained from humanized NSG mice before (Pre-AMG 191) and 2 weeks posttreatment of AMG 191 (1.0 mg/kg; Post-AMG 191). (B) Frequency of human hematopoietic progenitor cells (hCD45⁺hCD34⁺hCD117⁺) among all live cells present in BM aspirates obtained from humanized NSG mice treated with 0.3 mg/kg (left panel) and 1.0 mg/kg AMG 191 (right panel). P values were obtained using the paired Student t test. (C) A second human donor cell engraftment is facilitated by AMG 191 conditioning. Pooled cord blood CD34⁺ cells transduced with mCitrine-expressing lentivirus were transplanted into unconditioned and AMG 191–treated humanized NSG mice (0.3 and 1.0 mg/kg, see supplemental Materials and methods for details). Second donor grafts were infused on days 21 and 25 post–AMG 191 administration, days when the PK level was predicted to fall below 2000 ng/mL (supplemental Figure 5). The choice of this threshold level was based on PK studies in mice that showed that serum levels <2200 ng/mL of the anti-mouse CD117 antibody had no effect on donor mouse HSC engraftment (supplemental Figure 6). Six weeks after transplantation, chimerism was accessed in BM of transplanted humanized NSG mice. Secondary engraftment was measured by the frequency of mCitrine-expressing cells in each cell subset. Cell frequency was analyzed by FACS and FlowJo software. P values were obtained using the unpaired Student t test and Prism software. Data and error bars in panels B and C represent the mean plus or minus standard error of the mean (sem).